Etude fonctionnelle de la protéine prion cellulaire dans deux populations de cellules immunitaires

Les Encéphalopathies Spongiformes Subaiguës Transmissibles (ESST) sont des pathologies neurodégénératives qui présentent des déterminismes multiples. On distingue ainsi, les ESST familiales, sporadiques et infectieuses. Quelque soit leur forme, ces maladies sont généralement associées à une protéine appelée protéine du prion ou PrP. Notamment dans les ESST infectieuses, un homologue conformationnel de cette protéine est suspecté d être le constituant principal de l agent infectieux. Les deux formes de PrP sont identifiées sous les termes de cellulaire (PrPC), pour la protéine native et de scrapie (PrPSc), pour son homologue pathologique. Beaucoup d incertitudes planent sur la physiopathologie associée aux ESST, notamment parce que la fonction physiologique de la PrPC n est pas connue précisément. La protéine prion cellulaire est une glycoprotéine, ancrée via un groupement GlycosylPhosphatidylInositol, dans le feuillet externe de la membrane. La fonction physiologique de la PrP a été abondamment étudiée et la protéine a été impliquée dans de nombreux mécanismes cellulaires comme la transduction de signaux, la modulation de l apoptose, l adhésion cellulaire ou la reconnaissance des bactéries du genre Brucella. Cependant, beaucoup de ces études sont controversées et aucune ne fait l unanimité. Nous nous sommes intéressés, à notre tour, à la fonction physiologique de la PrPC dans deux modèles de cellules immunitaires : les lymphocytes Tg9 2 humains, sur lesquels nous avons observé une expression membranaire de la protéine très abondante et les macrophages murins, qui ont été impliqués dans la reconnaissance et la survie intracellulaire des Brucella. Nous avons étudié l éventuelle participation de la PrPC dans les divers mécanismes physiologiques associés à ces lymphocytes, d une part et à l infection des macrophages par Brucella, d autre part. Or, les résultats obtenus n ont pas permis de dégager une fonction physiologique évidente de la PrPC dans ces cellulesPrion diseases are neurodegeneratives pathologies of multiples determinisms. Inherited, sporadic or infectious forms are all associated with the expression of one protein, called prion protein or PrP. Infectious diseases are suspected to be caused by an abnormal conformational isoform of the cellular prion protein (PrPC) named scrapie or PrPSc . Physiopathologic events associated with prion diseases are not completely understood, in part because physiological function of PrPC is not precisely known. PrPC is a glycoprotein located in the extracellular side of the membrane with GPI anchorage. Due to its location, PrPC has been implicated in many cellular functions as signalling, apoptosis modulation, cellular adhesion or receptor for the bacteria Brucella. However, many of these functions are not clear or are controversial. We try to determine the physiological role of cellular prion protein by studying different cellular mechanisms of two populations of immune cells : Tg9 2 lymphocytes in which important expression of PrP has been found and murine macrophages where PrP has been implicated in the entry and intracellular proliferation of Brucella. But, in both cases, we were unable to demonstrate a functional implication of the cellular prion proteinMONTPELLIER-BU Sciences (341722106) / SudocSudocFranceF

Physiological role of the cellular prion protein

The prion protein (PrP) plays a key role in the pathogenesis of prion diseases. However, the normal function of the protein remains unclear. The cellular isoform (PrP$^{C})$ is expressed most abundantly in the brain, but has also been detected in other non-neuronal tissues as diverse as lymphoid cells, lung, heart, kidney, gastrointestinal tract, muscle, and mammary glands. Cell biological studies of PrP contribute to our understanding of PrP$^{\rm C}$ function. Like other membrane proteins, PrP$^{\rm C}$ is post-translationally processed in the endoplasmic reticulum and Golgi on its way to the cell surface after synthesis. Cell surface PrP$^{\rm C}$ constitutively cycles between the plasma membrane and early endosomes via a clathrin-dependent mechanism, a pathway consistent with a suggested role for PrP$^{\rm C}$ in cellular trafficking of copper ions. Although PrP$^{-/-}$ mice have been reported to have only minor alterations in immune function, PrP$^{\rm C}$ is up-regulated in T cell activation and may be expressed at higher levels by specialized classes of lymphocytes. Furthermore, antibody cross-linking of surface PrP$^{\rm C}$ modulates T cell activation and leads to rearrangements of lipid raft constituents and increased phosphorylation of signaling proteins. These findings appear to indicate an important but, as yet, ill-defined role in T cell function. Recent work has suggested that PrP$^{\rm C}$ is required for self-renewal of haematopoietic stem cells. PrP$^{\rm C}$ is highly expressed in the central nervous system, and since this is the major site of prion pathology, most interest has focused on defining the role of PrP$^{\rm C}$ in neurones. Although PrP$^{-/-}$ mice have a grossly normal neurological phenotype, even when neuronal PrP$^{\rm C}$ is knocked out postnatally, they do have subtle abnormalities in synaptic transmission, hippocampal morphology, circadian rhythms, and cognition and seizure threshold. Other postulated neuronal roles for PrP$^{\rm C}$ include copper-binding, as an anti- and conversely, pro-apoptotic protein, as a signaling molecule, and in supporting neuronal morphology and adhesion. The prion protein may also function as a metal binding protein such as copper, yielding cellular antioxidant capacity suggesting a role in the oxidative stress homeostasis. Finally, recent observations on the role of PrP$^{\rm C}$ in long-term memory open a challenging field

Absence of Evidence for the Participation of the Macrophage Cellular Prion Protein in Infection with Brucella suis

Brucella spp. are stealthy bacteria that enter host cells without major perturbation. The molecular mechanism involved is still poorly understood, although numerous studies have been published on this subject. Recently, it was reported that Brucella abortus utilizes cellular prion protein (PrP(C)) to enter the cells and to reach its replicative niche. The molecular mechanisms involved were not clearly defined, prompting us to analyze this process using blocking antibodies against PrP(C). However, the behavior of Brucella during cellular infection under these conditions was not modified. In a next step, the behavior of Brucella in macrophages lacking the prion gene and the infection of mice knocked out for the prion gene were studied. We observed no difference from results obtained with the wild-type control. Although some contacts between PrP(C) and Brucella were observed on the surface of the cells by using confocal microscopy, we could not show that Brucella specifically bound recombinant PrP(C). Therefore, we concluded from our results that prion protein (PrP(C)) was not involved in Brucella infection

A Micellar On-Pathway Intermediate Step Explains the Kinetics of Prion Amyloid Formation

International audienceIn a previous work by Alvarez-Martinez et al. (2011), the authors pointed out some fallacies in the mainstream interpretation of the prion amyloid formation. It appeared necessary to propose an original hypothesis able to reconcile the in vitro data with the predictions of a mathematical model describing the problem. Here, a model is developed accordingly with the hypothesis that an intermediate on-pathway leads to the conformation of the prion protein into an amyloid competent isoform thanks to a structure, called micelles, formed from hydrodynamic interaction. The authors also compare data to the prediction of their model and propose a new hypothesis for the formation of infectious prion amyloids

Old Gray Mouse Lemur Behavior, Cognition, and Neuropathology

International audienceNonhuman primate models are required to understand aging and age-related pathologies. The gray mouse lemur Microcebus murinus, a small prosimian primate, develops age-dependent deficits that are comparable to the decline observed during normal and pathological aging in humans. Importantly, not all old gray mouse lemurs are equally affected by age-related behavioral and cognitive problems. Some are profoundly impaired, while others perform as well as younger animals. Moreover, brain atrophy is detected only in some animals and thus appears to be an age-related pathological condition more than an inevitable effect of age. Finally, a subset of aged animals display neuropathological lesions observed also in Alzheimer's disease: β-amyloid deposition mainly in diffuse plaques and tau protein aggregation in some pyramidal neurons of the entorhinal cortex and hippocampus. Overall, these age-related changes indicate that gray mouse lemurs could be used as a potential translational model to study age-associated deficits and disorders

Dynamics of polymerization shed light on the mechanisms that lead to multiple amyloid structures of the prion protein.

International audienceIt is generally accepted that spongiform encephalopathies result from the aggregation into amyloid of a ubiquitous protein, the so-called prion protein. As a consequence, the dynamics of amyloid formation should explain the characteristics of the prion diseases: infectivity as well as sporadic and genetic occurrence, long incubation time, species barriers and strain specificities. The success of this amyloid hypothesis is due to the good qualitative agreement of this hypothesis with the observations. However, a number of difficulties appeared when comparing quantitatively the in vitro experimental results with the theoretical models, suggesting that some differences should hide important discrepancies. We used well defined quantitative models to analyze the experimental results obtained by in vitro polymerization of the recombinant hamster prion protein. Although the dynamics of polymerization resembles a simple nucleus-dependent fibrillogenesis, neither the initial concentration dependence nor off-pathway hypothesis fit with experimental results. Furthermore, seeded polymerization starts after a long time delay suggesting the existence of a specific mechanism that takes place before nucleus formation. On the other hand, polymerization dynamics reveals a highly stochastic mechanism, the origin of which appears to be caused by nucleation heterogeneity. Moreover, the specific structures generated during nucleation are maintained during successive seeding although a clear improvement of the dynamics parameters (polymerization rate and lag time) is observed. We propose that an additional on-pathway reaction takes place before nucleation and it is responsible for the heterogeneity of structures produced during prion protein polymerization in vitro. These amyloid structures behave like prion strains. A model is proposed to explain the genesis of heterogeneity among prion amyloid